O148 Shiga toxin-producing Escherichia coli outbreak: microbiological investigation as a useful complement to epidemiological investigation.

An outbreak of Shiga toxin-producing Escherichia coli (STEC) O148 infection occurred among wedding attendees in France in June 2002. A retrospective cohort study was performed and ten cases were identified, including two adults with haemolytic uraemic syndrome (HUS). The analytical study revealed that > 80% of affected individuals had eaten lightly roasted mutton and poultry pâté, but only the consumption of pâté tended to be associated with illness (relative risk 3.4; 95% CI 0.8-14.4). Left-overs (cooked mutton and raw offal) and processed foods (pâté) from the same batches as served at the party were sampled. Human, food and environmental samples were examined for the Shiga toxin (stx) gene and virulence traits by PCR. Stx-positive samples were cultured for STEC. HUS cases were tested for serum antibodies against 26 major STEC serogroups. An STEC O26 strain (stx1, eae, ehxA) was isolated from one case with diarrhoea, and an STEC O148 strain (stx2c) from one case of HUS. Serum antibodies against O26 were not detected in either of these patients; antibodies against O148 were not tested. Three STEC strains were isolated from the mutton and the offal (stx2c, O148), and two from the pâté (stx2c, O-X and O-Y). The isolates from the mutton were indistinguishable from the human stx2c isolate, whereas the pâté isolates differed. Although four different STEC strains were identified in patients and foods, the results of molecular subtyping, in conjunction with analysis of food consumption patterns, strongly suggested that this outbreak was caused by mutton contaminated with STEC O148.

Comparison of 14 PCR systems for the detection and subtyping of stx genes in Shiga-toxin-producing Escherichia coli.

The specificity of 14 polymerase chain reaction (PCR) systems designed for the detection and subtyping of stx genes was tested on a set of Escherichia coli strains with known sequences of stx genes. Systems designed for the detection of genes of the stx1 type did not detect any variant genes of the stx2 type and conversely, no stx2 type-specific systems detected stx1 variant genes. Among five stx2 type-specific systems, none detected the stx2ev gene, and two detected the stx2e gene. Among systems designed for screening genes of the both stx1 and stx2 types with a single primer pair, only one system (the Lin system) was able to detect stx genes in all studied strains. Shiga-toxin-producing E. coli frequently carry more than one stx variant gene. Coamplification of stx genes present in the same strain was demonstrated by restriction of PCR products with endonucleases generating fragments of variant-specific size. The amplification product obtained by the Lin system restricted by Hincll yielded fragments of different size for stx1, stx2, stx2c, stx2e and stx2ev. Thus it was possible to identify different genes carried in a single strain with a simple two-step PCR/endonuclease restriction protocol.

Molecular characterization of Vibrio cholerae O1 strains isolated in Romania.

A collection of 89 Vibrio cholerae O1 strains, isolated in Romania between 1977 and 1994, and 6 strains from the Republic of Moldavia, was characterized by ribotyping, toxin gene restriction pattern (toxinogenotype) and distribution of cholera toxin gene (ctx), accessory toxin gene (ace) and zonula occludens toxin gene (zot). After Bg/I endonuclease restriction of chromosomal DNA, a total of 18 ribotypes and 21 toxinogenotypes were distinguished. Deletions in the core region of the toxin gene cassette were found in 20% of strains; however, with the exception of one strain, all the isolates contained the ctx gene. Used in association, the three methods of molecular typing provided an accurate characterization of V. cholerae O1 isolates.